Summary of Work: In order to better understand the role of DNA replication fidelity in mutagenesis, we have isolated strains of E. coli that replicate their DNA with increased fidelity (antimutator mutants). They were obtained as suppressors of the high mutability of strains defective in postreplicative mismatch repair (mutL) or in exonucleolytic proofreading (mutD). In virtually all cases, the responsible mutation was found to reside in the dnaE gene, encoding the alpha subunit of DNA polymerase III responsible for replicating the bacterial chromosome. In addition to lowering the mutation rate in mutL strains by 3- to 20-fold, they were capable of lowering the mutation rate in an otherwise wild-type background by about 2-fold. This suggests that in E. coli uncorrected DNA replication errors may represent about one half of all spontaneous mutations. DNA sequencing of lacI forward mutations revealed that this one half is almost entirely composed of transversion base-pair substitutions, suggesting that these represent uncorrected replication errors. In contrast, the transitions may result from unrepaired DNA damages. We have also investigated the possible mechanisms by which the new DNA polymerase alleles increase the fidelity of replication. The experiments, both in vivo and in vitro, suggest that the antimutator dnaE alleles do not exert their effect directly via increased base selection, but indirectly by either slowing extension from mispaired bases (allowing increased proofreading) or by promoting polymerase dissociation from mispaired termini (allowing error removal by alternative means). We are currently investigating whether the increased accuracy of DNA replication occurs equally during the leading and lagging strand strand replication or whether it may be specific to one of them.